tbuckley/cabot-search huggingface dataset:
Abstract P2060: Gja1-20k Expression Increases With Left Ventricular Mechanical Unloading In Patients With Arrhythmogenic Cardiomyopathy And Advanced Heart Failure
Authors: Lindsey A Taylor, Steven Valdez, Christos Kyriakopoulos, JING LING, Craig Selzman, William Caine, Stavros G Drakos, Robin M Shaw, Joseph Palatinus
Journal: Circulation Research (2023)

GJA1-20k, the internally translated isoform of the gap junction protein Connexin-43 (Cx43) is required for trafficking of the full length Cx43 to the membrane. We recently found that GJA1-20k expression is reduced in patients and mice with arrhythmogenic cardiomyopathy (ACM), and that gene therapy-mediated introduction of GJA1-20k in a mouse model of ACM rescues Cx43 trafficking to the intercalated disc and reduces arrhythmia burden, indicating that loss of GJA1-20k contributes to arrhythmogenesis in ACM. Here, we examined cardiac muscle from patients with a confirmed diagnosis of ACM for GJA1-20k expression before and after left ventricular assist device (LVAD) support. The goal of this study was to determine if left ventricular (LV) unloading benefits endogenous expression of this critical trafficking protein. Cardiac tissue was collected from consecutive, prospectively enrolled patients undergoing LVAD implantation or heart transplantation in hospitals comprising the Utah Cardiac Recovery program and donor patients from hospitals throughout the Intermountain West. Samples were prepared for analysis using a standard lysis buffer and analyzed via Western Blotting using antibodies to GAPDH and Cx43-CT. Consistent with prior findings, cardiac muscle samples from patients with ACM had decreased GJA1-20k expression relative to muscle from healthy donor hearts not allocated for transplantation. However, tissue obtained from the same ACM patients post LVAD support (at the time of heart transplantation) revealed a large and significant increase in GJA1-20k expression compared to the pre LVAD timepoint (1 +/- 0.5 vs 10.2+/- 1, p=0.0013, n=3 ACM pre/post). In contrast to patients with ACM, patients with non-ischemic cardiomyopathy (NICM) that underwent LVAD implantation had no significant change in GJA1-20k expression after LV unloading (1+/- 0.5 vs 1.4+/- 0.7, p= 0.424, n=4 NICM pre/post). These data indicate that LV mechanical unloading increases GJA1-20k expression in hearts of patients with known ACM. Further work will focus on how mechanical unloading regulates GJA1-20k expression and whether mechanical unloading is sufficient to recover Cx43 trafficking and limit arrhythmias in ACM.
Abstract P3200: SH3BGR Affects Energy Metabolism In Neonatal Rat Cardiomyocytes
Authors: Anushka Deshpande, Hermine Lehmann, Norbert Frey, Ashraf Rangrez
Journal: Circulation Research (2023)

Background: The SH3 domain-binding glutamic acid-rich (SH3BGR) gene maps to the genetic fragment associated with Down's syndrome (DS), one of the predominant causes of congenital heart diseases (CHD). We have previously shown that SH3BGR augments cardiomyocyte apoptosis, however, its possible role in cardiomyocyte metabolism is not known yet. Methods: Functional effects of SH3BGR knockdown or overexpression were studied in neonatal rat ventricular cardiomyocytes (NRVCMs) using seahorse metabolic assays, quantitative real-time PCR, immunoblotting, and immunofluorescence microscopy. Results: Since we found significantly altered cardiac levels of SH3BGR in human patients suffering from cardiac hypertrophy, which is reportedly associated with mitochondrial dysfunction, we determined if loss- or gain-of-function of SH3BGR impacts mitochondrial metabolism. We observed significant reduction in oxidative phosphorylation complexes and associated genes. Interestingly, seahorse metabolic activity assays revealed impairment in mitochondrial bioenergetics by both knockdown and overexpression of SH3BGR suggesting the importance of its stead-state levels in cardiomyocyte mitochondrial metabolism. NRVCMs showed inability to meet high energy demand on altered SH3BGR expression indicated by higher proton leak and impeded ATP production. These observations suggest altered mitochondrial respiration, which was further supported by alterations in the expression levels of respective metabolic genes. Combining these results with our previous findings suggests that SH3BGR plays an important role in cardiomyocyte oxidative phosphorylation, cell viability and apoptosis. Conclusion: Our data provide novel insights into potential role of SH3BGR in cardiomyocyte mitochondrial bioenergetics. These findings will help us understand how genetic deletion or SH3BGR mutations are associated with CHD e.g., using CRISPR/CAS mediated genetic manipulations using human patient derived induced pluripotent stem cells.
Abstract P3176: One Carbon Metabolism Defect In Failing Heart
Authors: Jae Woo W Jung, Emily Flam, Kenneth Bedi, Cholsoon Jang, Joshua D Rabinowitz, Kenneth B Margulies, Zoltan Arany
Journal: Circulation Research (2023)

Aberrant cardiac metabolism has long been hypothesized to contribute to heart failure (HF). However, understanding of cardiac metabolism in HF remains incomplete. To fill this gap, we performed metabolomic and RNA sequencing analysis of 48 non-failing (NF) and 39 dilated cardiomyopathy (DCM) hearts. Among many differences, metabolites from one-carbon metabolism were significantly reduced, especially methionine (Met) and s-adenosylmethionine (SAM), a universal methyl donor of methylation. We probed the mechanism behind decreased Met/SAM in DCM hearts. Studies in neonatal rat ventricular myocytes (NRVMs), and in vivo steady-state infusions with isotope tracing showed that Met was mostly imported, rather than recycled, suggesting a defective Met transport in DCM hearts. We next explored the impact of low SAM. DNA and histone methylation remained unchanged, but creatine level was significantly decreased. So, we tested potential creatine synthesis in the heart that was previously unclear. To our surprise, NRVMs synthesized ~50% of their creatine pool when external supply is low, and synthesis was dependent on SAM availability. In summary, we find that the majority of Met in cardiomyocyte is imported, with rapid turnover. We also find that cardiomyocytes synthesize creatine and increase creatine production under stress. Together, these findings suggest that defective Met import may contribute to low creatine and SAM levels and ultimately HF.
Abstract P2073: Hyperbaric Oxygen Therapy Protects The Myocardium From Reductive Stress-induced Proteotoxic Remodeling
Authors: Sini Sunny, jennifer Houg, Shibu Navaneeth, Sayed Aniqa, Afortude John Kofi, Rajasekaran N Namakkal-Soorappan
Journal: Circulation Research (2023)

Background: Oxygen is the prime source of reactive oxygen species (ROS) in cells. We hypothesize that hyperbaric oxygen therapy (HBOT) can be used as an effective intervention to maintain redox homeostasis in the myocardium. Here, we assessed whether HBOT could prevent RS through enhanced ROS signaling, thereby protecting the hearts from proteotoxic stress. Methods: Cardiac-specific human Nrf2 transgenic (hNrf2-TG) and their non-transgenic littermates (NTG) were used in this study. Mice at the age of 8 weeks were exposed to hyperbaric oxygen (2.5 Bars, 2 hr/day) for 8 weeks. The TG and NTG (n=6) mice without HBOT served as controls. Pre- and post- echocardiography (Vevo 2100) analysis were carried out to assess the structural and functional changes over time in response to HBOT exposure. The influence of HBOT on matrixins (MMP2/9), redox (GSH &amp; NQO1), and cardiac hypertrophy markers (ANF, BNF) were examined using zymography, immunoblotting, and qPCR analysis. The aggregation of proteins in myocardial tissues was detected by Proteostat staining. Results: While the expression of hNrf2 induces RS in the mouse heart, we noticed that HBOT ameliorates the RS through enhanced ROS signaling. A decrease in systemic GSH levels along with the downregulation of ANF and BNF mRNA levels observed in the HBOT group indicates the restoration of the redox homeostasis as well as reduced remodeling of the myocardium. Interestingly, hNRF2-induced RS was suppressed by HBOT as supported by changes in the protein levels for NQO1, GCLC, and GCLM. The degree of protein aggregation in RS hearts was significantly reduced by HBOT. Vevo strain analysis reveals that 8 weeks of HBOT restored the diastolic stretch seen in the RS hearts to a normal diastole-systole velocity equal to NTG mice. A significant decrease in the ejection fraction (80% vs. 55% in TG vs. TG-HBOT), was evident in the TG mice that received HBOT. Repression of matrixins, observed in the hearts of HBOT-TG mice signifies the benefits of HBOT in RS hearts. Conclusion: HBOT directly improves myocardial structure at the tisuue level; maintain the redox status at systemic level, and improves the myocardial performance at functional level. Our study established that HBOT can rescue the myocardium from RS-induced proteotoxic insults.
Abstract P2078: Vasohibin Inhibition Improves Myocardial Relaxation In A Rodent Model Of Heart Failure With Preserved Ejection Fraction
Authors: Deborah M Eaton, Benjamin W Lee, Matthew A Caporizzo, Yingxian Chen, Amit Iyengar, Danika Meldrum, Jessica Dominic, Noah Weingarten, Khaled Hached, Kenneth Bedi, Pavan Atluri, Siem Van Der Laan, Benjamin L Prosser, Kenneth B Margulies
Journal: Circulation Research (2023)

Heart failure with preserved ejection (HFpEF) is a complex syndrome with few therapies available to patients. Diastolic dysfunction is a common clinical feature among HFpEF patients, and strategies to mitigate impaired relaxation may have a therapeutic benefit. Recognizing that increased stabilization of the microtubule network induced by detyrosination of α-tubulin contributes to increased cardiomyocyte stiffness ex vivo , we explored if inhibition of detyrosination with a small molecule vasohibin inhibitor (VASHi) could improve in vivo diastolic function in obese/hypertensive (ob) ZSF1 rats, compared to lean/hypertensive ZSF1 and non-hypertensive Wistar Kyoto (Wky) rats as controls. Animals received two intravenous injections of VASHi (6.6mg/kg) or vehicle (veh) at 30 weeks old, when ob have a HFpEF phenotype. Echocardiography and invasive hemodynamics were performed after the first and second injection, respectively. Compared to ob veh-treated animals, ob VASHi-treated animals had a significant improvement in invasively measured time constant of early LV relaxation, tau (veh: 14.73ms ± 0.73 vs. VASHi: 11.02ms ± 0.45; P &lt; 0.05) and echo-derived deceleration time of early LV filling, indicative of improved early relaxation (Fig. A, B). VASHi improved early relaxation in cardiomyocytes isolated from non-failing (NF, N=3) and failing (HF, N=4) human hearts (Fig. C), providing the first evidence of small molecule VASHi improving human myocyte function. These studies indicate that excess detyrosination contributes to impaired early myocyte and myocardial relaxation in a clinically relevant HFpEF model and supports a potential therapeutic role for VASHi in HFpEF.
Abstract P3067: Emerging Immune Checkpoint Therapies Reshape The Cardiac Immune Landscape And Promote Myocardial Vulnerability
Authors: Pan Ma, Ana Villanueva, Jing Liu, Takeshi Egawa, Chieh-Yu Lin, Javid Moslehi, Kory J Lavine
Journal: Circulation Research (2023)

Immune checkpoint therapies have shown remarkable efficacy across multiple tumor types by amplifying cancer-immunity and boosting T-cell immune response against tumors. As an increasing number of immune checkpoint molecules have been identified, numerous combination therapeutic strategies are under clinical investigation. For instance, the combination of an OX40 (TNFRSF4) agonist and PD-1 (Programmed cell death protein 1) inhibitor has been shown to have promising anti-tumor effects in pre-clinical studies and early clinical trials. Despite remarkable therapeutic benefit, it has become evident with increasing clinical usage that these agents elevate the risk of immune-related adverse events (irAEs), especially cardiovascular sequelae. Mitigating irAEs is critical for the establishment of efficacious and safe interventions. Currently, little is understood regarding the cardiac effects of combined OX40 activation and PD-1 inhibition. In this study, we observed that PD-1 deletion upregulated OX40 expression on cardiac T-cells. Combined PD1 inhibition (Pdcd1 -/- or PD-1 blocking antibody) and OX40 activation (OX40 agonistic antibody) increased the frequency of splenic and cardiac effector memory CD4 + and CD8 + T-cells, inflammatory monocyte and macrophage infiltration into the heart, and enhanced myocardial expression of numerous chemokines and cytokines in both naïve and tumor bearing mice. Intriguingly, persistent increases in cardiac effector memory CD4 + and CD8 + T-cells and inflammatory macrophages was observed at chronic time points suggesting long term effects on the cardiac immune landscape. Furthermore, OX40 activation aggravates angiotensin II and phenylephrine (AngII/PE) infusion induced cardiac hypertrophy, fibrosis and inflammation in Pdcd1 -/- mice suggesting that the reshaped T-cell and macrophage landscape imparts myocardial vulnerability. This study will also uncover mechanism-based therapeutic targets to prevent or treat cardiac adverse events elicited by combined OX40 activation and PD-1 inhibition.
Abstract P1069: Yap Mediates Cardiac Proliferation In Tead-independent Mechanisms
Authors: Bing Xie, Jeff Steimle, Vaibhav Deshmukh, Yuka Morikawa, Fansen Meng, James F Martin
Journal: Circulation Research (2023)

Myocardial infarction (MI) is a leading cause of death worldwide. During an MI, the contractile myocardium experiences reduced blood flow causing expansive cell death. Since adult mammalian hearts lack regenerative capacity, MI damage is permanent followed by heart failure. However, recent studies have demonstrated that the neonatal mouse heart exhibits transient regenerative potential within the first 7 days, suggesting a regulatory mechanism that prevents the adult from initiating a regenerative response to cardiac injury. The Hippo signaling pathway negatively regulates mammalian cardiac regeneration by inhibiting the activity of downstream effector YAP. However, YAP5SA, which completely bypasses Hippo pathway regulation, results in YAP hyper-transcriptional activity and heart overgrowth. In this work, we aim to investigate the TEAD-independent mechanisms of YAP in promoting cardiomyocyte (CM) division and renewal. We generated another YAP protein, which can not only skip Hippo pathway inhibition, but also lose the interaction with TEADs. We found that this modified YAP promoted CM proliferation and improved cardiac repair post MI injury. Our work will uncover novel functions of YAP and improve YAP therapeutics in myocardial infarction, finally benefiting human heart health.
Abstract P2070: The Microprotein Another-Regulin Regulates Serca2a In Response To Neurohormonal Signaling
Authors: Keira Hassel, Aaron Gibson, Cat A Makarewich
Journal: Circulation Research (2023)

Objective: Calcium (Ca 2+ ) regulation is a key component of proper heart function and is largely controlled by the sarco/endoplsmic reticulum Ca 2+ ATPase 2a (SERCA2a). Ca 2+ dysregulation is a hallmark feature of heart failure, yet few advances have been made to reestablish Ca 2+ homeostasis during disease. Another-regulin (ALN) inhibits SERCA2a, though little is known about how ALN functions in heart physiology and disease. Here, we determine a mechanism by which ALN is regulated via phosphorylation at serine 19 (S19), resulting in dissociation from SERCA2a. Methods and Results: ALN phosphorylation at S19 was confirmed via Phos-tag gel migration assay and site directed mutagenesis (SDM), whereby serine to alanine (S19A) mutation abolished the migrational shift which was confirmed with a custom S19 phospho-specific antibody. Effects of S19 phosphorylation on ALN-SERCA2a affinity were investigated using co-immunoprecipitation (IP) and fluorescence resonance energy transfer (FRET) in HEK293 cells. In addition to phosphodeficient S19A, SDM was used to mutate S19 to phosphomimetic aspartate (S19D) and glutamate (S19E). IPs of SERCA2a and hemagglutinin (HA)-tagged ALN WT, S19A, S19D, or S19E revealed less interaction between SERCA2a and ALN-S19D/E, suggesting S19 phosphorylation causes ALN dissociation from SERCA2a. Acceptor sensitization and fluorescence lifetime measurements show that ALN-S19D decreases ALN/SERCA2a binding affinity. To investigate signaling upstream of ALN phosphorylation, adult mouse cardiomyocytes were stimulated with various agonists and levels of pALN measured. Phosphorylation was significantly induced with the neurohormonal stimuli endothelin-1 and phenylephrine compared to DMSO and the β-adrenergic stimulus isoproterenol, suggesting Gαq/PKC pathway involvement. In addition, pALN levels are elevated in whole heart lysates of mice subject to transverse aortic constriction which is a model of pathological hypertension. Conclusions: These results indicate that ALN is phosphorylated at S19 in response to neurohormonal stimuli, resulting in dissociation from SERCA2. Future directions will investigate the impact of ALN phosphorylation on SERCA2 activity and heart function.
Abstract P2088: Proteostatic Imbalance In Heart Failure With Preserved Ejection Fraction Is Accompanied By Mechanistic Target Of Rapamycin Complex 1 Hyperactivation
Authors: Kamil A Kobak, Weronika Zarzycka, Catherine J King, Frederick F Peelor, Benjamin F Miller, Ying Ann Chiao
Journal: Circulation Research (2023)

Heart failure with preserved ejection fraction (HFpEF) is a multi-organ syndrome that represents approximately 50% of all heart failure (HF) cases. The limited effective therapies for HFpEF and poor understanding of the underlying mechanisms make this disorder a major challenge for cardiovascular medicine. Disruption of protein homeostasis (proteostasis) is a well-known pathomechanism involved in many chronic diseases and aging. However, how proteostasis changes during HFpEF development remains unclear. The objective of this study is to determine the role of proteostatic regulation in HFpEF pathology. We used a combination of metabolic and hypertensive stress (2-hit stress) to induce HFpEF development in C57BL/6 mice. Our proteomic comparison between HFpEF mice and unstressed controls revealed changes in the abundance of proteins involved in protein translation, protein folding, and the ubiquitin-proteasome system, suggesting that proteostatic maintenance is altered during HFpEF. Using deuterium oxide (D 2 O) labeling coupled with GC-MS analysis of subcellular fractions, we found higher protein synthesis rates in myofibrillar and mitochondrial fractions of HFpEF hearts compared to controls. Subsequent isotopic analysis of RNA synthesis showed that ribosomal biogenesis was 2-fold higher in HFpEF hearts compared to controls. We also observed a reduced LC3-II/I ratio in HFpEF hearts, which may suggest suppressed autophagy. These results indicate that HFpEF hearts have increased protein synthesis and reduced protein degradation compared to controls. As the mechanistic target of rapamycin complex 1 (mTORC1) is a master regulator of proteostasis that promotes protein synthesis and suppresses autophagy, we investigated if mTORC1 signaling is altered in HFpEF hearts. We found that HFpEF hearts had increased activation of mTORC1, as indicated by increased phosphorylation of its targets. In conclusion, we showed that HFpEF pathology is associated with proteostatic imbalance, potentially due to mTORC1 hyperactivation. Our ongoing study is investigating if mTORC1 inhibition (using cardiac-specific Raptor heterozygous knockout mice) preserves proteostatic maintenance and attenuates cardiac dysfunction during HFpEF development.
Abstract P3065: Mechanistic Insights Into Cardiac Dysfunction And Remodeling Associated With Invasive Pneumococcal Pathogenesis
Authors: Tousif Sultan, Daniel Minassian, Prachi Umbarkar, Chao He, Min Xie, Palaniappan Sethu, Carlos J Orihuela, Hind Lal
Journal: Circulation Research (2023)

Background: One-in-four adults hospitalized for community-acquired pneumonia (CAP) experiences a major adverse cardiac event (MACE). Pneumonia survivors, in particular those who experienced invasive pneumococcal disease, are also at elevated risk for MACE in convalescence. Although Streptococcus pneumoniae ( Spn )-mediated cardiac dysfunction is a well-established clinical problem, the mechanisms by which pneumonia impacts the long-term risk of MACE are poorly understood. Methods and Results: Each mouse was infected with 1 x 10 4 CFU of Spn , to model invasive pneumococcal disease (IPD). At 30 hours post-infection, mice were administered Ampicillin (50mg/kg) every 12 hours for 3 days. After that, animals were given Ampicillin (1mg/ml) ad libitum in drinking water for 4 days. Three weeks post-infection phase, Spn -rescued animals’ demonstrated marked cardiac dysfunction as reflected by significantly reduced ejection fraction (EF), and fractional shortening (FS). Cardiac injury was confirmed to have occurred by measurement of increased expression of ANP, BNP (q-PCR), and levels of serum Troponin-I as measured by ELISA. Trichrome-stained heart sections demonstrated fibrotic remodeling in the Spn -rescued animals. To gain insight into mechanisms, we analyzed whether Spn alters cardiac immune cell infiltration and pro-inflammatory S100A8/A9-TLR-NLRP3-IL-1β signaling axis in vivo . Indeed, Spn -rescued (with antibiotic) animals showed an increased percentage of TLR-2/4+, NLRP3+, and phosphor-NFkB+ myeloid/neutrophils in the heart. Importantly, these findings were corroborated using human blood samples collected from patients with IPD. Pharmacological blocking of S100A8/A9-TLR-NLRP3-IL-1β signaling axis by S100A9 inhibitor largely reduced Spn -induced detrimental cardiac pathologies in convalescence. Conclusion: Our results suggest chronic activation of the pro-inflammatory S100A8/A9-TLR2/4-NLRP3-IL-1β pathway in Spn -infected hearts that persists even after antibiotic clearance of bacterial infection. Targeting the S100A8/9-NLRP3-IL-1β signaling circuit reduced chronic inflammation and fibrosis in our mice model and therefore is potentially a novel strategy to mitigate the Spn-mediated cardiac dysfunction.